This document outlines a new genetic diagnosis protocol using whole exome capture and massively parallel DNA sequencing. The protocol allows for decreased DNA sequencing and cost while increasing detection of heterozygous bases and read depth. It was used to diagnose a patient with congenital chloride diarrhea through an unexpected genetic finding that confirmed the clinical diagnosis. The protocol has implications for discovering disease genes involved in Mendelian traits through identifying de novo mutations or homozygous segments, and for complex traits by applying selective sequencing of promising genes identified through initial sequencing of subjects.

1. Genetic diagnosis by whole
exome capture and massively
parallel DNA sequencing
Abdul Jaleel Yusif
Muhammed Emre Sevük
Mustafa Salih Oğuz

2. Outline
● Abstract
● Materials and Methods
● Results
● Discussion

3. Abstract

4. 85% of mutations with
large effects on
disease-related traits

5. Target
-D652N mutation at SLC26A3 (the known congenital chloride
diarrhea locus)
- Some patients who has no mutations at SLC26A3

6. Materials and Methods

7. Human Subjects
-Patients with Bartter syndrome(BS)
-consanguineous kindred subjects
with no mutation in genes causing BS.

10. Sonication for Shearing

12. Results
(a lot of tables and figures)

13. What are these?
● SNV
● Missense
● Allele
● Homozygous/Heterozygou
s

16. autosomal recessive inheritance

23. Discussion

24. Implications of the new Protocol
● Decrease in the amount of DNA sequencing
● Decrease in the cost of detection of exonic mutations
● High read depth, increase in detection of heterozygous bases (97.2%)
● Increased read per lane in the future

25. Clinical Implications
● New method: Unexpected genetic diagnosis (with an undiagnosed illness) +
Clinical diagnosis
-Congenital Chloride Diarrhea
-Mutation in SLC26A3
-Confirmation
● Parsing large amount of genetic data to produce clinically useful information using
the new method plus hints from clinical condition to arrive at a correct diagnosis.

26. Application
Discovering disease related genes and alleles connected with:
● Mendelian traits
● Complex traits
● Somatic mutations in cancers

27. Mendelian traits
❖ Dominant traits (Alleles that have been difficult to map via linkage analysis):
➢ Traits with substantial locus heterogeneity
➢ Solution: Identifying significant excess no of independent mutations in the
same gene by mapping data that constrains the location of the disease locus.
➢ Alleles that impair reproductive fitness such that affected subjects harbor De
Novo mutations.
➢ Solution: Finding the individual De Novo mutations will provide evidence of
disease

28. Mendelian traits
❖ Recessive Traits (Diseases due to consanguineous union):
➢ Disease locus is homozygous within a segment that is homozygous by descent
from a recent ancestor. Information search is therefore constrained to 10% of
the exome for first cousins.
➢ HNV detection in such segments has high efficiency even at low levels of per-
base coverage (5X).
➢ Number of protein-altering variants in these segments is very low (~29)
➢ Solution: KO in animal models (loss of function)

29. Complex Traits
❖ Non mendelian: derived from multiple genes and exhibit variety of phenotypes
➢ Sample size for loci identification is very large
➢ Apply new method for all subjects or a subset, then apply selective
sequencing for most promising genes.
❖ Missing some non-coding regions with rare variant that have large effects becomes
inevitable.

30. Conclusion
Whole-exome sequencing will make broad contributions to understanding the genes
and pathways that contribute to rare and common human diseases as well as clinical
practice.

31. Thank you for
listening!